In Vitro Contraction of Isolated Cauda Epididymal Duct Smooth Muscle as a Complimentary Approach to Physiological, Pathological, Toxicological, and Pharmacological Studies on Epididymal Function.

Contraction of cauda epididymal duct (CE) smooth muscle is one of the very first events of the seminal emission phase of ejaculation. The contraction of CE smooth muscle is governed by a complex interaction of hormones, autacoids, and by the neurotransmitters released from the epididymal intramural nerve endings, and any impairment in the CE smooth muscle contraction has the potential to impair male fertility. Apart the obvious pathophysiological and toxicological importance of CE smooth muscle contraction, modulation of CE contraction has pharmaceutical interest offering a druggable target to development of drugs to improve/impair male fertility. The in vitro contraction experiments constitute a valuable approach to an in-depth evaluation of functional and molecular changes resulting from pathologies or drug exposure. Therefore, this chapter consists in a description of in vitro pharmacological reactivity contractility of the epididymal duct in a controlled medium, maintained at 30 °C of temperature and continuously bubbled with 95% O2 and 5% CO2 to obtain cumulative concentration-response curves that has been fundamental to some of our investigations on epididymal physiology, toxicology, and pharmacology.

Impact of timing of the anorexigen sibutramine administration on reproductive end-points of male rats.

Sibutramine is a non-selective serotonin-norepinephrine reuptake inhibitor orally administered for weight loss. In a previous study, we showed pharmacological mechanisms involved in the reduction of sperm quality and fertility of rats exposed for 30 days to this anorexigen in the light phase of the light-dark (l/d) cycle. It is already known that rodents are nightlife animals, with higher metabolic activity during the dark phase than in the light phase of the light-dark (l/d) cycle. Thus, the present study aimed to investigate whether the deleterious effects on reproductive parameters after sibutramine administration would be enhanced after a shorter period of exposure during the dark phase of the l/d cycle. For this, adult male Wistar rats were treated with sibutramine (10 mg/kg/d) or vehicle for 15 days during the dark phase of the l/d cycle. Sibutramine treatment decreased final body and reproductive organ weights, as well as serum testosterone levels. Sperm transit time through the epididymis was accelerated, and sperm concentration and motility were diminished in the sibutramine-exposed rats. The decrease in sperm concentration was also verified in the epididymal histological sections. In conclusion, the deleterious effects of sibutramine on reproductive parameters of male rats were enhanced when the exposure occurred in the dark phase of the l/d cycle, even after a short exposure duration. Our results reinforce the impact of timing on drug therapeutic action.

Short- and long-term effects on reproductive parameters of female Wistar rats after exposure to rosuvastatin starting in pre-puberty.

Statins are a class of drugs that act lowering lipid levels by inhibiting cholesterol biosynthesis. Additionally, statins can act by "pleiotropic effects", related to the inhibition of synthesis of the other mevalonate pathway products. Rosuvastatin is a third-generation statin and has shown better results in reducing cholesterol concentrations when compared to other statins. Recent studies suggest that rosuvastatin may act as an endocrine disruptor that potentially damages the hormonal axis and, consequently reproductive development and function of male rats. However, the effects of rosuvastatin exposure on rat female reproductive parameters remain unknown. In this study female rats were exposed to rosuvastatin at the doses of 0 (control), 3, or 10 mg/Kg.bw-1/day from pre-puberty to adulthood. No alterations in the female reproductive parameters were observed at a dose of 3 mg/Kg.bw-1. However, females exposed to 10 mg/Kg.bw-1 exhibited shorter estrous cycles, altered copulatory behavior, decreased serum prolactin level, and alterations in the liver, pituitary and placental weights, parameters to some extent influenced by the reproductive hormonal axis signaling pathway. On the other hand, pubertal onset, reproductive hormone levels, fertility, and histological parameters of the ovary, uterus, and placenta were unaltered by exposure to both doses of this statin. Thus, rosuvastatin exposure, at the higher dose, altered the reproductive function of female rats, probably due to the pleiotropic effects of this statin. Additional studies on the effects of this statin on female reproductive function and development are encouraged to better characterize its mode of action.

Reproductive disorders in female rats after prenatal exposure to betamethasone.

Betamethasone is the drug of choice for antenatal treatment, promoting fetal lung maturation and decreasing mortality. Previous studies in rats reported male programming and alteration in sperm parameters and sexual behavior following intrauterine betamethasone exposure. The impact on the female reproductive development is not known. In this study, rat female offspring was assessed for sexual development, morphophysiology of the reproductive tract and fertility after maternal exposure to 0.1 mg kg-1 of betamethasone or vehicle on gestational days 12, 13, 18 and 19. The treatment promoted reduction of litter weight on postnatal day 1, morphological masculinization in females, delay in the age of puberty onset, reduction in estrus number, increase in estrous cycle length and increase in luteinizing hormone serum levels and uterus weight. The females from the betamethasone group showed an increase of myometrial uterine area and decrease in endometrial uterine area. These animals also performed less lordosis during the sexual behavior test and showed impaired reproductive performance. The uterus showed higher contraction in the treated group as shown by a pharmacological assay. In conclusion, prenatal betamethasone exposure in rats promoted female masculinization, altered sexual development and reproductive parameters. Copyright © 2017 John Wiley & Sons, Ltd.

Perinatal exposure to insecticide fipronil: effects on the reproductive system in male rats.

Fipronil is an insecticide widely used in agriculture, veterinary medicine and public health that has recently been listed as a potential endocrine disrupter. In the present study we evaluated the effects of perinatal exposure to fipronil during the period of sexual brain differentiation and its later repercussions on reproductive parameters in male rats. Pregnant rats were exposed (via gavage) to fipronil (0.03, 0.3 or 3mgkg-1) from Gestational Day 15 until Postnatal Day 7. Fipronil exposure did not compromise the onset of puberty. In adulthood, there was no effect on organ weight or sperm production. Furthermore, there were no adverse effects on the number of Sertoli cells per seminiferous tubule, testicular and epididymal histomorphometry or histopathology or expression patterns of androgen receptor in the testis. Similarly, no changes were observed in the sexual behaviour or hormone levels. However, in rats exposed to fipronil, changes in sperm motility were observed, with a decrease in motile spermatozoa and an increase in non-mobile spermatozoa, which can compromise sperm quality in these rats. Perinatal exposure to fipronil has long-term effects on sperm parameters, and the epididymis can be a target organ. Additional studies should be undertaken to identify the mechanisms by which fipronil affects sperm motility.

Maternal exposure to butyl paraben impairs testicular structure and sperm quality on male rats.

Parabens are hormonally active chemicals widely used as preservatives in foods and are frequently detected in human fluids and tissues. Therefore, the objective of this study was to determine the effects of maternal butyl paraben (BP) exposure on male sexual development. Pregnant Wistar rats received corn oil (control group), or BP at doses of 10, 100, or 200 mg/kg, subcutaneously, from gestational day 12 until postnatal day 21. Our results demonstrated that developmental BP exposure significantly increased the number of adult Leydig cells and the circulating concentrations of testosterone and attenuated FSH and LH concentrations at 200 mg/kg. BP exposure adversely affected spermatogenesis kinetics at doses of 10 and 200 mg/kg and provoked a decrease in the immunostaining of EsR1 and AR at 200 mg/kg. The sperm motility was impaired at the 10 mg/kg dose, and sperm head abnormalities were increased in all BP dose groups. We suggest that BP impairs testicular structure and function in the rat, affecting sperm quality. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1273-1289, 2017.

Alterations in male rats following in utero exposure to betamethasone suggests changes in reproductive programming.

Antenatal betamethasone is used for accelerating fetal lung maturation for women at risk of preterm birth. Altered sperm parameters were reported in adult rats after intrauterine exposure to betamethasone. In this study, male rat offspring were assessed for reproductive development after dam exposure to betamethasone (0.1mg/kg) or vehicle on Days 12, 13, 18 and 19 of pregnancy. The treatment resulted in reduction in the offspring body weight, delay in preputial separation, decreased seminal vesicle weight, testosterone levels and fertility, and increased testicular weight. In the testis, morphologically abnormal seminiferous tubules were observed, characterized by an irregular cell distribution with Sertoli cell that were displaced towards the tubular lumen. These cells expressed both Connexin 43 (Cx43) and Proliferative Nuclear Cell Antigen (PCNA). In conclusion, intrauterine betamethasone treatment appears to promote reproductive programming and impairment of rat sexual development and fertility due to, at least in part, unusual testicular disorders.

Slimmer or fertile? Pharmacological mechanisms involved in reduced sperm quality and fertility in rats exposed to the anorexigen sibutramine.

Sperm acquire motility and fertility capacity during epididymal transit, under the control of androgens and sympathetic innervations. It is already known that the acceleration of epididymal sperm transit time can lead to lower sperm quality. In a previous work we showed that rats exposed to the anorexigen sibutramine, a non-selective serotonin-norepinephrine reuptake inhibitor, presented faster sperm transit time, lower epididymal sperm reserves and potentiation of the tension of epididymal duct to norepinephrine exposed acutely in vitro to sibutramine. In the present work we aimed to further investigate pharmacological mechanisms involved in these alterations and the impact on rat sperm quality. For this, adult male Wistar rats were treated with sibutramine (10 mg/kg/day) or vehicle for 30 days. Sibutramine decreased final body, seminal vesicle, ventral prostate and epididymal weights, as well as sperm transit time in the epididymal cauda. On the contrary of the in vitro pharmacological assays, in which sibutramine was added directly to the bath containing strips of distal epididymal cauda, the ductal tension was not altered after in vivo sub-chronic exposure to sibutramine. However, there is pharmacological evidence that the endogenous epididymal norepinephrine reserves were reduced in these animals. It was also shown that the decrease in prostate weight can be related to increased tension developed of the gland, due to sibutramine sympathomimetic effects. In addition, our results showed reduced sperm quality after in utero artificial insemination, a more sensitive procedure to assess fertility in rodents. The epididymal norepinephrine depletion exerted by sibutramine, associated with decreases in sperm transit time, quantity and quality, leading to reduced fertility in this experimental model, reinforces the concerns about the possible impact on fertility of man taking sibutramine as well as other non-selective serotonin-norepinephrine reuptake inhibitors, especially considering the lower reproductive efficiency of humans compared to males of other species.